Hepatitis C virus (HCV) infection poses a major global health problem, infecting approximately 4 million people in the United States and 170 million people worldwide. The properties and biochemical composition of HCV virions have not been determined due largely to the lack of a robust culture system for HCV infection and propagation. Recently, we have successfully established a stable culture system for robust production of infectious HCV. The characterization of HCV RNA-containing particles revealed that the low-density HCV particles were of the highest infectivity in sharp contrast to the higher-density particles, which contained the highest level of HCV RNA genome but were not infectious. Additionally, apolipoproteins B and E were detected in the low- density fractions of HCV particles, and HCV infection was potently inhibited by human lipoproteins LDL and VLDL. Therefore, we hypothesize that HCV virion is assembled with lipoproteins, which play important roles in the mediation of HCV infection through specific interactions with cell surface receptors/co-receptors. HCV virion assembly with lipoproteins represents a novel concept that warrants exploratory investigation and holds promise for discovering novel antiviral drugs. The overall goal of this application is to determine the properties and biochemical composition of HCV RNA-containing particles and to define the role of lipoproteins in HCV assembly and infection. Our specific aims are: 1) to determine the nature and biochemical composition of HCV RNA-containing particles and the structural determinants of HCV infectivity using structural, cell biological, biochemical, immunological, and proteomic approaches; 2) to define the role of lipoproteins in HCV virion production and infection by altering the levels of lipoprotein production in stable HCV-producing cells as well as by using antibodies against specific apolipoproteins. The levels of lipoproteins will be either up regulated by apolipoprotein-expression vectors and chemical inducers or down regulated by specific siRNAs and small molecular inhibitors. Additionally, we will determine the underlying mechanisms for the inhibition of HCV infection by human lipoproteins. Findings derived from these studies will not only determine the molecular basis underlying HCV infectivity but also provide novel targets for antiviral intervention. Furthermore, information gained from our studies will provide a foundation towards a thorough understanding of the molecular mechanisms of HCV virion assembly and infection. Hepatitis C virus (HCV) infection poses a major global health problem, infecting about 4 million people in the United States and 170 million people worldwide. This application is to determine the properties and biochemical composition of HCV particles and to define the role of lipoproteins in HCV production and infection. Findings derived from these studies will provide novel targets and therapies for antiviral intervention of HCV infection. [unreadable] [unreadable] [unreadable]